Hello,
I am trying to induce my BL21 cells with IPTG and I will purify my his tagged- protein using Ni-NTA column. I am using a lysis buffer of 25mM Tris-Cl, 50mM glucose and 10mM EDTA pH8.0 and adding lysozyme to this before starting. What I am not using is any kind of protease inhibitor. I wanted to add PMSF in my lysis buffer as well but I am not sure If it is ok or it may cause a problem with lysozyme in terms of activities. EDTA is not recommended for example but I couldn't find anything for PMSF...
Does anybody know if it is ok to add these two together ?
Thank you!
It is fine to add PMSF to inhibit proteases, it won't interfere with lysozyme. Be advised that PMSF powder is not soluble in water, you have to make a stock solution in methanol and it is poisonous, so careful when handling it. Now a day there a protease inhibitor cocktails that work well as Complete from Roche, you might want have a look at them.
Yes you are right about that. I wa using PMSF without lysozyme this is the first time I want to combine them so I couldn't be sure. Roche cocktail we have I guess but I thought PMSF was better for bacteria. I may be wrong but I can give a trial. Thank you.
u can use pmsf, it wont interact with lysozyme. but i suggest better to use protease inhibitor cocktail.
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